The work of the laboratory for this year was centered on four aspects of the effects of ketone body metabolism. Manufacture and Use of Ketone Body Monoesters All the ketone body monoesters, D--hydroxybutyrate R 1,3 butanediol monoester, used in all the studies reported here were made in this laboratory by Todd King using methods devised here. The purity of the ester and its optical properties were analyzed by Dr Robert Pawlosky using chromatographic resolution principles and mass spec methods devised here. Further tests of the pharmacokinetics of these ketones esters in human subjects were performed by our colleagues at Oxford (1). The ketone esters have GRAS status and all use in humans complied with protocol requirements approved by the Oxford University IRB. The list of 10 collaborators to which we have supplied ketone ester is appended to this report. Ketone Body Metabolism Enhances Physiological and Cognitive Performance The relationship of redox states to phosphorylation potential was described in 1969 (Krebs HA &Veech RL, Bari Symposium) However it required 10 years to determine the precise relationship of the energy of the phosphorylation potential required determination of the intracellular free Mg2+ (Veloso D et al JBC 248: 4811-119, 1973) and the resultant changes in the equilibrium constants of phosphate containing metabolic reactions to be able to determine the energy of the hydrolysis of ATP , the central reaction of energy metabolism ( Veech RL and Krebs, HA 1979 JBC 254: 6538-47) . The ability of ketone body metabolism to overcome insulin resistance and the increase in the G of ATP hydrolysis was first determined in working perfused heart (Sato, N et al FASEB J 9: 651-58, 1995). This year we extended this work to human subjects where Dr Clarke and her group showed that the metabolism of ketone esters improved the physical performance in elite athletes (2) thus fulfilling our obligation to DARPA which originally provided the funds for our work on ketone bodies. This paper was published in Cell Metabolism and was cited as among the best publications in that journal for 2016. In a second paper in this area, done in collaboration with workers from both Cambridge and Oxford, it was shown that rats fed a ketone ester diet showed enhanced cognitive performance (3) Ketone Bodies Alter the Localization and Activity of Transcription Factors In collaboration with investigators at Southwestern U Medical school, it was demonstrated in hepatic cells that the ketone body D--hydroxybutyrate altered the nuclear localization of the transcription factor carbohydrate-response element binding protein, ChREBP, thereby inhibiting fat synthesis (4). In a later section of this report, another effect of D--hydroxybutyrate on transcription is described as an inhibitor of histone deacetylase thus increasing the activity of antioxidant enzymes. The Metabolism of Ketone Esters By Passes the Loss of Insulin Sensitivity. Causing Alzheimers Disease By Replenishing A Deficiency of Krebs Cycle Metabolites The description and review of the causes, resulting from a loss of insulin activation of cerebral pyruvate dehydrogenase and the treatment of resultant Alzheimers disease with ketone bodies is presented (5). Comparing samples from the frontal cortex to those from hippocampal regions from the brains of 3XTgAD mice chronically fed a ketone ester diet, Dr Pawlosky and his collaborators showed (6) that the deficiency in Krebs cycle intermediates in the hippocampal region was reversed in this model of Alzheimers disease. The importance of these findings toward the development of a therapeutic strategy for the treatment of Alzheimers disease was the topic of a special article in the Journal of Neurochemistry (Puchowicz M.A. et al, J Neurochem. 141: 162-4, 2017). The Metabolism of Ketone Bodies Reduces the Free NADP+ /NADPH Ratio Thus Mitigating Oxygen Free Radical Damage In 1969 we had shown that the free NADP+ /NADPH ratio, with a redox potential of -0.42V was the most negative in the body (Krebs,H.A and Veech RL, 1969). In 1995, we showed that the metabolism of ketone bodies reduce the NADP couple. We now reviewed the use of redox active agents, showing why the use of single redox active agents such as ascorbic acid and the like were without demonstrable in vivo effects but that the metabolism of ketone bodies due to the linkage of the intracellular redox couple could mitigate oxygen free radical damage (7). The ability of ketone bodies to mitigate the damage of ionizing radiation from rogue nuclear devices has been presented to the Nuclear Bioterrorism and Training Group, the Nuclear Radiation Commission, the Interagency Steering Committee on Radiation Standards and the NIAID Countermeasures Program. Ketone Bodies Mimic the Life Span Extending Properties of Caloric Restriction. Caloric restriction has long been known to extend life span in many species from nematodes to mice. Feeding ketones can decrease signaling in the insulin/ insulin grow factor receptor, a major factor in the genetic pathway of life span extension. D--hydroxybutyrate is the natural inhibitor of the histone deacetylase which represses the transcription of the FOXO gene thus increasing the transcription of antioxidant enzymes. Thus ketosis decreases insulin signaling, increases antioxidant enzymes and increases the reducing potential to diminish ROS damage which plays an important role in prolonging life span (8)